Clinical and microbiological characteristics of and risk factors for bloodstream infections among patients with extracorporeal membrane oxygenation: a single-center retrospective cohort study

Extracorporeal membrane oxygenation (ECMO) provides hemodynamic and oxygenation support to critically ill patients. Due to multiple catheter cannulations, patients on ECMO are vulnerable to bloodstream infections (BSIs). We aimed to investigate the incidence, clinical characteristics, risk factors, and microorganisms associated with BSIs during ECMO. This single-center retrospective cohort study was conducted between January 2015 and May 2021. Patients aged 18 years or older with an ECMO duration of > 48 h for cardiogenic or respiratory support were included in the study. Patients who developed bacteremia or candidemia from 12 h after ECMO cannulation to 7 days after de-cannulation were included. The clinical factors between non-BSI and BSI were compared, along with an analysis of the risk factors associated with BSI during ECMO. A total of 480 patients underwent ECMO for cardiogenic shock (n = 267, 55.6%) or respiratory failure (n = 213, 44.4%) during the study period. The incidence was 20.0 episodes per 1000 ECMO-days. Approximately 20.2% (97/480) and 5.4% (26/480) of the patients developed bacteremia and candidemia, respectively. The median numbers of days of BSI development were 8.00 days for bacteremia and 11.0 days for candidemia. The most common pathogens were methicillin-resistant coagulase-negative staphylococci (n = 24), followed by vancomycin-resistant Enterococcus (n = 21). Multivariable logistic analysis demonstrated that hemodialysis (odds ratio [OR] 2.647, p < 0.001), veno-arterial-venous mode (OR 1.911, p = 0.030), and total ECMO duration (OR 1.030, p = 0.007) were significant risk factors for bacteremia. The total ECMO duration was the only risk factor associated with candidemia (OR 1.035, p = 0.010). The mortality rate was significantly higher in the bacteremia (57.7%) and candidemia (69.2%) groups than that in the non-BSI group (43.6%). BSI is a common complication of patients receiving ECMO support and is associated with poor clinical outcomes. Determining the type of frequently isolated organisms and the median onset time of BSI would help in the selection of appropriate prophylactic antibiotics or antifungal agents.


Methods
Patient selection. In this single-center, retrospective, observational cohort study, adult patients who underwent ECMO at Severance Hospital between January 2015 and May 2021 were initially screened (n = 769). Severance Hospital is a 2400-bed tertiary referral hospital that performs various lung and heart surgical procedures, including organ transplantations. Patients who met the following criteria were included in the study: (1) aged 18 years or older, (2) who received ECMO support beyond 48 h, and (3) who received ECMO for the first time. Patients who received repeated ECMO were excluded from the analysis. This study was approved by the institutional review board of Severance Hospital (No. 4-2021-1587). The requirement for informed consent was waived owing to the retrospective nature of the study. All procedures and treatments observed current guidelines and regulations. Data collection. Patients' clinical information, demographics, and laboratory data at the initiation of ECMO were obtained from the Severance Clinical Research Analysis Portal. The data included age, sex, body mass index, underlying medical conditions, and medications. Detailed information related to ECMO was obtained from a daily ECMO report written by physicians. This information included the reasons for ECMO support (cardiogenic vs. respiratory failure), mode of ECMO (veno-arterial [VA], veno-venous [VV], or veno-arterialvenous [VAV]), cannulation site of ECMO catheters, location of catheter cannulation (intensive care unit, operating room, emergency room, or intervention room), type of ECMO machine (prolonged life support [PLS] or emergency bypass system [EBS]), and total time of ECMO implantation. Additionally, data on the application of renal replacement therapy were also collected. Data on antibiotic usage, blood culture results during ECMO support, type of organisms isolated, and antibiotic sensitivity test results were collected. The antibiotics administered prior to or during ECMO cannulation were analyzed in this study. ECMO procedure. Intensive care specialists, cardiologists, or cardiothoracic surgeons determined the need for ECMO initiation by evaluating the patient's medical condition. The ECMO techniques used in our institution have been described in a previously published study 7 . Cannulation was performed using the Seldinger technique, and the preferred cannulation sites were the femoral and internal jugular veins for VV ECMO and the femoral vein and artery for VA ECMO. If additional cardiogenic support was required for patients receiving VV ECMO, an additional arterial catheter was inserted and the mode was switched to VAV ECMO. For the analysis, the specific mode with the highest number of catheters inserted was determined for each ECMO mode. All patients who underwent ECMO cannulation were infused with heparin (2000 IU). The target activated coagulation time was 180-200 s. The Capiox EBS (Terumo Co., Ltd., Tokyo, Japan) or Bioline heparin-coated Quadrox PLS circuit system (Maquet Cardiopulmonary, Hirrlingen, Germany) was used in all patients. All ECMO patients requiring addition of continuous renal replacement therapy (CRRT) used separate circuits for each system.  (Fig. 1). Table 1 displays the baseline characteristics of patients in the non-BSI and BSI groups (bacteremia and candidemia group). The mean age of the participants was 56.3 ± 14.5 years, and 65.4% of them were men. Patients in the BSI group were older than those in the non-BSI group (58.8 vs. 55.5, p = 0.024). The BSI group had a lower proportion of patients with congestive heart failure (CHF) (42.9% vs. 53.5% vs. p = 0.048); moreover, the candidemia group had a lower proportion of patients with CHF than in the non-BSI group (30.8% vs. 53.5% www.nature.com/scientificreports/ p = 0.026). The number of patients requiring hemodialysis, including intermittent or continuous, was higher in the BSI group (59.8% vs. 41.1%, p = 0.001), especially in the bacteremia group. The median length of pre-ECMO hospital stay was significantly longer in the bacteremia (23.0 days) and candidemia (25.5 days) groups than in the non-BSI group (13.0 days). Over 70% of the patients were already receiving antibiotics, and more than 60% were using antibiotics with Pseudomonas coverage. The rate of antibiotic use before ECMO did not differ between the non-BSI and BSI groups.

Duration of ECMO and risk of BSI.
The total duration of ECMO support in median days was higher in the bacteremia and candidemia groups (13.0 and 17.2 days) compared with that in the non-BSI group (6.11 days) (p = 0.003 and 0.001, respectively). The in-hospital mortality rate was also higher in the bacteremia and candidemia groups (57.7% and 69.2%, respectively) compared with that in the non-BSI group (43.6%) (p = 0.013 and 0.012, respectively). The Kaplan-Meier curves in Fig. 2a,b show that the mortality curve rapidly dropped in both the bacteremia and candidemia groups compared with that in the non-BSI group. Risk factor associated with BSI in ECMO. The risk factors associated with bacteremia and candidemia during ECMO were analyzed using univariable and multivariable logistic regression analyses ( Table 2). In univariable analysis, age, hemodialysis, respiratory support, VAV mode, ECMO system with PLS, cannulation procedure performed in the intervention room, pre-ECMO hospital days, and total duration of ECMO were the risk factors of bacteremia. In the multivariable analysis, only hemodialysis (OR 2.647, 95% CI 1.547-4.528, p < 0.001), VAV mode (OR 1.911, 95% CI 1.065-3.429, p = 0.030), and total duration of ECMO (OR 1.030, 95% CI 1.008-1.053, p = 0.007) were the significant clinical factors associated with an increased risk of bacteremia. Moreover, only the total duration of ECMO (OR 1.035, 95% CI 1.008-1.061, p = 0.010) was associated with the risk of candidemia.
Major causal pathogens of BSI during ECMO. Table 3 lists the frequently isolated causal pathogens of bacteremia or candidemia during ECMO along with the first isolated time of each organism (in median days). Among the gram-negative organisms, Acinetobacter baumannii and Klebsiella pneumoniae were the most commonly detected pathogens of BSI during ECMO. Both organisms displayed multidrug resistance to antibiotics. All isolated Escherichia coli was either extended-spectrum beta-lactamase-producing (5 out of 6) or multidrugresistant organism (1 out of 6). Coagulase-negative staphylococcus (methicillin resistant) and enterococcus (vancomycin resistant) were common pathogens in gram-positive organisms. Candida tropicalis was the most common Candida species, followed by Candida parasilosis and Candida albicans. Figure 4 shows the microorganisms isolated on the first median days, in chronological order, to visualize the pathogens responsible for developing BSI in the early or later phase of ECMO. Bacteremia due to Enterobacter cloacae and Escherichia coli occurs in the earlier phase (less than 7 days) after ECMO cannulation. By contrast, vancomycin-resistant enterococcus, Candida glabrata, Candida parasilosis, and Morganella morganii were causal pathogens of BSI during ECMO, which developed in the later phase (after 15 days) of ECMO cannulation. Table 4 shows the cases in which the causal pathogens were isolated from other sources before causing BSI. Furthermore, the causative microorganisms in BSI 37 out of 98 (37.8%) patients in the bacteremia group and 9 out of 26 (34.6%) patients in the candidemia group were previously isolated from other sources, such as sputum, urine, bronchial alveolar lavage, pleural drainage catheter, swab culture, endotracheal tube, or stool. Approximately 25.5% and 23.1% of bacteremia and candidemia pathogens, respectively, were isolated after ECMO cannulation. The most commonly isolated sources were sputum and bronchial alveolar lavage for bacteremia and sputum and urine for candidemia.

Discussion
BSI is a critical complication of ECMO since BSI during ECMO is associated with increased mortality and morbidity 2 . The BSI rate during ECMO was higher in adults than in neonates and children 10 . Therefore, preventing the occurrence of BSI in adults is crucial. However, there are insufficient data to guide the active surveillance or the selection of prophylactic antibiotics.
The incidence of BSI during ECMO (23.3%) reported in this study was higher than the previously reported incidence of 3-18% 2,11,12 . This high incidence might be attributed to our extended definition of BSI during ECMO, which included BSI occurring up until 7 days after de-cannulation. Since the skin barrier is broken during ECMO cannulation and requires sufficient time to heal depending on the patient's medical condition after catheter removal, the incidence of BSI was evaluated for an extended period.
The significant risk factors associated with bacteremia during ECMO were hemodialysis, VAV mode, and total duration of ECMO. The hemodialysis and VAV modes may be related to the need for additional indwelling www.nature.com/scientificreports/ catheters. Many ECMO patients require renal replacement therapy during ECMO for acute kidney injury (AKI) due to hemodynamic instability. To provide hemodialysis (HD) or CRRT, a permanent catheter or a temporary catheter is required. In this study, we did not determine whether the risk of BSI differs in patients requiring hemodialysis due to end-stage renal disease (ESRD) or in those requiring hemodialysis due to AKI. A previous study reported a risk of BSI in patients with elevated creatinine levels 13 . Therefore, impaired renal function requiring hemodialysis is associated with a risk of BSI during ECMO. The critical risk factors associated with BSI development were extended hospital stay and total duration of ECMO. Multiple studies demonstrated that the total duration of ECMO was the most critical risk factor of Table 2. Clinical variables associated with bloodstream infection (bacteremia, candidemia). p values with statistical significance is shown in italics text. *Indicates bacteremia group. † Indicates candidemia group. BMI body-mass index, COPD chronic obstructive pulmonary disease, ECMO extracorporeal membrane oxygenation, VA veno-arterial, VV veno-venous, VAV veno-arterial-venous, PLS prolonged life support, EBS emergency bypass system.   [13][14][15] . The longer the patient stays in the ward or intensive care unit, the higher the risk of acquiring nosocomial pathogens. Therefore, shortening the total ECMO duration and hospital stay should be considered if the patient's hemodynamic stability is restored. Different ECMO modes were also associated with BSI during ECMO. The reason for ECMO support was used to determine the initial mode of ECMO. In Sun et al. 's study, the risk for using the VV mode was relatively higher (OR 4.473, CI 1.001-19.977, p = 0.050), probably because patients requiring respiratory support may have already developed infections prior to receiving ECMO support 6 . Other studies have also reported that most BSI cases were due to the use of respiratory ECMO 5,16 . VV ECMO was associated with BSI due to gram-negative bacteremia compared with the VA mode 16 . In this study, multivariable logistic analysis proved that VAV mode was associated with increased risk of developing bacteremia (OR 2.846, p < 0.001). Although not significant, the VV mode was also associated with an increased risk of developing bacteremia (OR 1.504, p = 0.153). As the VV mode is initially applied to patients with respiratory failure due to pneumonia, acute respiratory distress syndrome, or lung transplantation, such patients may have a higher risk of carrying the associated microorganisms in the lungs.
As the circuit regulates the body temperature of ECMO patients, clinicians often face difficulties in detecting the signs and symptoms of infection 17 . Leukocytosis and C-reactive protein levels are useful laboratory markers for evaluating the risk of infection in the absence of fever. In particular, if patients on ECMO support for longer than 10 days present with leukocytosis and low-grade fever, infection should be suspected 4 . Our study results showed that the median onset time of BSI was 8.0 days for bacteremia and 11.0 days for candidemia. Therefore, patients who demonstrate signs of infection beyond 8 or 11 days after ECMO cannulation, with elevated levels  www.nature.com/scientificreports/ of laboratory markers, could be considered for prophylactic antibiotic or antifungal treatment due to the possibility of infection, especially BSI. Taking the median onset time of BSI into account for each ECMO system, patients using VA, VV, and VAV ECMO longer than 7, 9, and 13.5 days, respectively, should be carefully assess the signs of infection. Consistent with the findings of previous studies, our study also noted CoNS as the most frequently isolated microorganism in BSI during ECMO 14 . Membrane oxygenators have large artificial contact surfaces with blood and are a potential source of infection during ECMO 2 . CoNS can bind to fibronectin, colonize membrane surfaces, and produce biofilms 18 . However, routine use of glycopeptides might not be helpful in ECMO patients. Clinicians often add glycopeptides when infections are suspected in patients on ECMO support. However, inappropriate antibiotic use increases the risk of acquiring multidrug resistance. According to our data, both the non-BSI and BSI groups used antibiotics including antipseudomonal agents and glycopeptides. However, the use of these antibiotics did not reduce the incidence of BSI. Approximately 42% of ECMO centers systematically administer antibiotics to all patients, with varying timings 19 . The administration of antibiotics without consideration of the possibility of infection has no benefits. Because most nosocomial infections develop after a median of 9.5 days, our data do not support the need for early antimicrobial prophylactic treatment, which may target drug-resistant pathogens. In other studies, prior exposure to carbapenem was associated with Candida BSI 16 .
Fungal prophylaxis may be helpful because Candida BSI is associated with higher mortality in ECMO patients than in the general population 19 . A previous study demonstrated that appropriate fungal prophylaxis in pediatric patients decreased the incidence of fungal infections 17 . Since the median onset time for candidemia is approximately 11 days after ECMO cannulation, an anti-fungal agent may be considered for patients who show signs of infection 11 days after ECMO insertion 2,20 .
Finally, this study emphasizes the importance of environmental control in hospitals. According to our data, most causal microorganisms were isolated from the respiratory samples. Previous studies have reported that the prevalence of hospital-acquired infection during ECMO is 10-12%, and multidrug-resistant pathogens were detected in 28.6% of patients 17,21 . With extended hospital stay, the risk of acquiring various microorganisms, including MDROs, is higher. In this study, several microorganisms isolated from blood cultures were subjected to MDROs. Knowing the commonly isolated microorganisms of the intensive care unit of the institution and the colonizing organism is essential for selecting the appropriate empirical antibiotics for treating BSI in ECMO patients.
Our study has some limitations. It was conducted in a single-center and was retrospective and observational in nature. Information on additional devices, such as intra-aortic balloon pumps, the use of distal perfusion catheter insertion, and a temporary pacemaker, may help evaluate the impact of BSI on ECMO. Complications such as AKI, distal perfusion failure, and skin necrosis might be other risk factors associated with BSI during ECMO support.

Conclusions
BSI is a common complication of ECMO and is associated with poor clinical outcomes. The risk factors associated with BSI in ECMO should be well established to improve the prognosis. Considering the type of frequently isolated organisms and the median onset time of BSI would help select the appropriate prophylactic antibiotics or antifungal agents.

Data availability
The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.